Methods for detecting assay interferents and increasing dynamic range
Inventors
Pope, Mark R. • Hershberger, Stefan J. • Ruan, Qiaoqiao • Swift, Kerry M. • Tetin, Sergey Y. • MacDonald, Patrick • Haack, Richard A.
Assignees
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Publication Number
US-12704519-B2
Publication Date
2026-08-11
Expiration Date
Abstract
The disclosure provides kits and methods for detecting a substance that interferes with detection of an analyte in a sample and for expanding the dynamic range and reducing the hook effect of an immunoassay. The kits and methods employ two conjugates with two different detectable labels, at least one of which is a chemiluminescent compound of Formula (I).
Core Innovation
The disclosure describes chemiluminescent reporter compounds and immunoassay kits using detectable-label conjugates and solid-support binding members to detect an analyte and interfering substances. Solid-support binding members bind the analyte and interfering substances concurrently or competitively, alongside optional support-bound members specific for the interferent. Across disclosed configurations, at least one detectable label comprises a compound of formula (I), or a salt thereof, based on an acridinium-fluorophore scaffold with shifted-wavelength emission.
Additional kit configurations employ two labeled binding members for the same analyte with differing binding affinities, or utilize different detectable labels. These configurations determine analyte concentration by evaluating dual-label signals against a flag value or using two-channel photomultiplier tube detection to expand dynamic range and reduce hook effects. Documented formula (I) conjugates include fluorescein, rhodamine, BODIPY, Alexa Fluor, silicon-rhodamine, Atto 700, and Lucifer Yellow derivatives.
Claims Coverage
The independent claims are claims 1, 5, 6, and 8, which recite four inventive features directed to immunoassay kits incorporating at least one detectable label comprising a compound of formula (I), or a salt thereof.
Kit for detecting an analyte and an interfering substance
A kit comprises a first conjugate whose binding member specifically binds an analyte, a second conjugate whose binding member specifically binds a substance interfering with analyte detection, and a solid-support binding member that binds the analyte and interfering substance concurrently or competitively, with an optional support-bound binding member specifically binding the interfering substance, wherein at least one detectable label comprises a compound of formula (I), or a salt thereof.
Kit for detecting interfering biotin
A kit comprises a biotin-bearing detectable-label binding member that specifically binds an analyte, a second detectable-label analyte-binding conjugate, and a streptavidin-coated solid support that binds both the biotin on the first binding member and biotin that interferes with analyte detection, wherein at least one detectable label comprises a compound of formula (I), or a salt thereof.
Kit with analyte-binding members of different affinities
A kit comprises two labeled binding members that bind the same analyte, wherein the first binding member exhibits greater binding affinity for the analyte than the second binding member, and a solid-support binding member that binds the analyte concurrently with either labeled binding member, wherein at least one detectable label comprises a compound of formula (I), or a salt thereof.
Kit with different detectable labels
A kit comprises a first conjugate and a second conjugate having different detectable labels and specific binding members that are the same or different, together with a solid-support binding member that binds the analyte concurrently with either conjugate, wherein at least one detectable label comprises a compound of formula (I), or a salt thereof.
The independent claims define four kit configurations directed to detecting assay interferences such as biotin, utilizing varying binding affinities, or using different detectable labels, wherein each kit requires at least one label comprising a compound of formula (I) or a salt thereof.
Stated Advantages
Expands immunoassay dynamic range.
Reduces the immunoassay hook effect.
Dual-channel assays detect and correct interference from anti-thyroglobulin antibody, anti-troponin antibody, and biotin, including signal recovery of NGAL under biotin interference.
Documented Applications
Dual-channel assay configurations for detecting and correcting interference from substances including autoantibodies, anti-thyroglobulin antibody, anti-troponin antibody, macroprolactin in a prolactin assay, and biotin, including NGAL signal recovery under biotin interference.
Determination of analyte concentration in immunoassays using signals associated with two detectable labels and a flag value to expand dynamic range and reduce hook effect, demonstrated with TSH.
Troponin signal correction using a measured relationship between interferent-channel signal and assay bias.
Immunoassay execution utilizing solid supports such as streptavidin-coated supports and spatially separated electrodes.
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